Human Sco1 and Sco2 function as copper-binding proteins

Yih Chern Horng, Scot C. Leary, Paul A. Cobine, Fiona B.J. Young, Graham N. George, Eric A. Shoubridge, Dennis R. Winge

Research output: Contribution to journalArticle

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Abstract

The function of human Sco1 and Sco2 is shown to be dependent on copper ion binding. Expression of soluble domains of human Sco1 and Sco2 either in bacteria or the yeast cytoplasm resulted in the recovery of copper-containing proteins. The metallation of human Sco1, but not Sco2, when expressed in the yeast cytoplasm is dependent on the co-expression of human Cox17. Two conserved cysteines and a histidyl residue, known to be important for both copper binding and in vivo function in yeast Sco1, are also critical for in vivo function of human Sco1 and Sco2. Human and yeast Sco proteins can bind either a single Cu(I) or Cu(II) ion. The Cu(II) site yields S-Cu(II) charge transfer transitions that are not bleached by weak reductants or chelators. The Cu(I) site exhibits trigonal geometry, whereas the Cu(II) site resembles a type II Cu(II) site with a higher coordination number. To identify additional potential ligands for the Cu(II) site, a series of mutant proteins with substitutions in conserved residues in the vicinity of the Cu(I) site were examined. Mutation of several conserved carboxylates did not alter either in vivo function or the presence of the Cu(II) chromophore. In contrast, replacement of Asp238 in human or yeast Sco1 abrogated the Cu(II) visible transitions and in yeast Sco1 attenuated Cu(II), but not Cu(I), binding. Both the mutant yeast and human proteins were nonfunctional, suggesting the importance of this aspartate for normal function. Taken together, these data suggest that both Cu(I) and Cu(II) binding are critical for normal Sco function.

Original languageEnglish
Pages (from-to)34113-34122
Number of pages10
JournalJournal of Biological Chemistry
Volume280
Issue number40
DOIs
Publication statusPublished - 2005 Oct 7

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Yeast
Yeasts
Copper
Fungal Proteins
Mutant Proteins
Ions
Cytoplasm
Reducing Agents
Chromophores
Chelating Agents
Aspartic Acid
Cysteine
copper-binding protein
Charge transfer
Bacteria
Proteins
Substitution reactions
Ligands
Recovery
Geometry

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Horng, Y. C., Leary, S. C., Cobine, P. A., Young, F. B. J., George, G. N., Shoubridge, E. A., & Winge, D. R. (2005). Human Sco1 and Sco2 function as copper-binding proteins. Journal of Biological Chemistry, 280(40), 34113-34122. https://doi.org/10.1074/jbc.M506801200
Horng, Yih Chern ; Leary, Scot C. ; Cobine, Paul A. ; Young, Fiona B.J. ; George, Graham N. ; Shoubridge, Eric A. ; Winge, Dennis R. / Human Sco1 and Sco2 function as copper-binding proteins. In: Journal of Biological Chemistry. 2005 ; Vol. 280, No. 40. pp. 34113-34122.
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Horng, YC, Leary, SC, Cobine, PA, Young, FBJ, George, GN, Shoubridge, EA & Winge, DR 2005, 'Human Sco1 and Sco2 function as copper-binding proteins', Journal of Biological Chemistry, vol. 280, no. 40, pp. 34113-34122. https://doi.org/10.1074/jbc.M506801200

Human Sco1 and Sco2 function as copper-binding proteins. / Horng, Yih Chern; Leary, Scot C.; Cobine, Paul A.; Young, Fiona B.J.; George, Graham N.; Shoubridge, Eric A.; Winge, Dennis R.

In: Journal of Biological Chemistry, Vol. 280, No. 40, 07.10.2005, p. 34113-34122.

Research output: Contribution to journalArticle

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AU - Horng, Yih Chern

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AU - Cobine, Paul A.

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AU - George, Graham N.

AU - Shoubridge, Eric A.

AU - Winge, Dennis R.

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AB - The function of human Sco1 and Sco2 is shown to be dependent on copper ion binding. Expression of soluble domains of human Sco1 and Sco2 either in bacteria or the yeast cytoplasm resulted in the recovery of copper-containing proteins. The metallation of human Sco1, but not Sco2, when expressed in the yeast cytoplasm is dependent on the co-expression of human Cox17. Two conserved cysteines and a histidyl residue, known to be important for both copper binding and in vivo function in yeast Sco1, are also critical for in vivo function of human Sco1 and Sco2. Human and yeast Sco proteins can bind either a single Cu(I) or Cu(II) ion. The Cu(II) site yields S-Cu(II) charge transfer transitions that are not bleached by weak reductants or chelators. The Cu(I) site exhibits trigonal geometry, whereas the Cu(II) site resembles a type II Cu(II) site with a higher coordination number. To identify additional potential ligands for the Cu(II) site, a series of mutant proteins with substitutions in conserved residues in the vicinity of the Cu(I) site were examined. Mutation of several conserved carboxylates did not alter either in vivo function or the presence of the Cu(II) chromophore. In contrast, replacement of Asp238 in human or yeast Sco1 abrogated the Cu(II) visible transitions and in yeast Sco1 attenuated Cu(II), but not Cu(I), binding. Both the mutant yeast and human proteins were nonfunctional, suggesting the importance of this aspartate for normal function. Taken together, these data suggest that both Cu(I) and Cu(II) binding are critical for normal Sco function.

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Horng YC, Leary SC, Cobine PA, Young FBJ, George GN, Shoubridge EA et al. Human Sco1 and Sco2 function as copper-binding proteins. Journal of Biological Chemistry. 2005 Oct 7;280(40):34113-34122. https://doi.org/10.1074/jbc.M506801200